Testing the efficacy of kitasamycin for use in the control and treatment of swine dysentery in experimentally infected pigs.

BACKGROUND: Swine dysentery (SD) caused by Brachyspira hyodysenteriae is an important disease in Australia. AIM: The aim of this study is to evaluate the macrolide antibiotic kitasamycin for use in SD control. METHODS: The minimum inhibitory concentrations (MICs) of kitasamycin, tylosin and lincomycin for 32 Australian isolates of B. hyodysenteriae were evaluated. Mutations in the 23S rRNA gene were examined. Isolate '13' with a low kitasamycin MIC was used to challenge weaner pigs. Sixty pigs were housed in 20 pens each containing three pigs: pigs in four pens received 2 kg/tonne of a product containing kitasamycin (3.1% active) prophylactically in their food starting 4 days before B. hyodysenteriae challenge (group 1); pigs in four pens were challenged and received the same dose therapeutically once one pig in a pen showed diarrhoea (group 2); four pens were challenged and received 4 kg/tonne of the product therapeutically (group 3); four pens were challenged but not medicated (group 4); two pens were unmedicated and unchallenged (group 5) and two pens received 2 kg/tonne and were unchallenged (group 6). Pigs were monitored for B. hyodysenteriae excretion and disease. RESULTS: Macrolide resistance was widespread, and mutations in the 23S rRNA gene were identified in 23 isolates. Four isolates with kitasamycin MICs < 5 µg/mL were considered susceptible. Following experimental challenge, 10 of 12 unmedicated pigs developed SD. No pigs receiving kitasamycin prophylactical or therapeutically developed SD. Medicated pigs shed low numbers of B. hyodysenteriae in their faeces. CONCLUSIONS: Kitasamycin can help control SD in pigs infected with susceptible isolates of B. hyodysenteriae.

Seeking the sun in deep, dark places: mesopelagic sightings of ocean sunfishes (Molidae).

Evidence is presented from publicly available remotely operated vehicle (ROV) footage that suggests deep-water ranging in ocean sunfishes (family Molidae) is more common than typically thought, including a new maximum depth recorded for the southern sunfish Mola ramsayi.

Identification of weakly haemolytic Brachyspira isolates recovered from pigs with diarrhoea in Spain and Portugal and comparison with results from other countries.

Weakly haemolytic anaerobic intestinal spirochaetes of the genus Brachyspira are commonly identified based on species-specific gene sequences. Apart from the pathogenic Brachyspira pilosicoli, the distribution and disease associations of the other weakly haemolytic Brachyspira species in pigs have not been comprehensively investigated. In this study weakly haemolytic Brachyspira isolates (n=67) from Spanish and Portuguese pigs with diarrhoea, negative in a routine diagnostic PCR for B. pilosicoli, were identified by sequencing their NADH oxidase genes (nox). Nearly half the isolates were identified as Brachyspira murdochii (n=31; 46.3%). The others were Brachyspira innocens (n=26; 38.8%), Brachyspira intermedia (n=7; 10.4%), "Brachyspira pulli" (n=1; 1.5%) and a potentially novel Brachyspira species (n=2; 3%). Multilocus sequence typing (MLST) on a subset of 18 isolates confirmed their species designations, including the potential new species, and identified similarities to strains from other countries.

Diets containing inulin but not lupins help to prevent swine dysentery in experimentally challenged pigs.

Swine dysentery is a contagious mucohemorrhagic diarrheal disease caused by the intestinal spirochete Brachyspira hyodysenteriae that colonizes and induces inflammation of the cecum and colon. It has been reported that a diet containing chicory root and sweet lupin can prevent swine dysentery. This experiment was conducted to test the hypothesis that inulin in the chicory root rather than galactans in lupins was responsible for protective effects. An experiment with a 2 × 2 factorial arrangement of treatments was undertaken using pigs fed barley- and triticale-based diets, with the main effects being protein source [185 g/kg of canola meal (decreased galactans) or 220 g/kg of lupins (greater galactans)] and inulin supplementation (0 or 80 g/kg). Forty Large White × Landrace pigs weighing 21 ± 3 kg, with 10 pigs per diet, were allowed to adapt to the diets for 2 wk, and then each pig was challenged orally 4 times with a broth culture containing B. hyodysenteriae on consecutive days. Pigs were killed when they showed clinical signs of dysentery or 6 wk postchallenge. Pigs fed diets without inulin had 8.3 times greater risk (P = 0.017) of developing swine dysentery and were 16 times more likely (P = 0.004) to have colon contents that were culture-positive for B. hyodysenteriae, compared with the pigs fed a diet with 80 g/kg of inulin. Diets containing lupins did not prevent pigs from developing clinical swine dysentery; however, inclusion of lupins or inulin or both in the diets delayed the onset of disease compared with the diet based mainly on canola meal (P < 0.05). Diet did not influence the total concentration of organic acids in the ileum, cecum, or upper and lower colon; however, the molar proportions of the organic acids were influenced (P < 0.05). Consequently the pH values in the cecum, and upper and lower colon were not influenced (P > 0.05) by diet. However the pH values of the ileal digesta were decreased in pigs fed the diet with both lupins and inulin compared with the diet containing only lupins (P < 0.05). In conclusion, this study shows that diets supplemented with highly fermentable carbohydrates from inulin protected pigs against developing swine dysentery.

Development of a multiplex-PCR for rapid detection of the enteric pathogens Lawsonia intracellularis, Brachyspira hyodysenteriae, and Brachyspira pilosicoli in porcine faeces.

AIMS: To develop an assay to simultaneously detect Lawsonia intracellularis, Brachyspira hyodysenteriae and Brachyspira pilosicoli in pig faeces. METHODS AND RESULTS: A multiplex-polymerase chain reaction (M-PCR) was designed to amplify a 655-base pair (bp) portion of the L. intracellularis 16S rRNA gene, a 354-bp portion of the B. hyodysenteriae NADH oxidase gene, and a 823-bp portion of the B. pilosicoli 16S rRNA gene. Specificity was assessed using 80 strains of Brachyspira spp. and 30 other enteric bacteria. Bacterial DNA was extracted from faeces using the QIAamp DNA Stool Mini Kit. The M-PCR was tested in parallel with culture and/or PCR on 192 faecal samples from eight piggeries. Faeces also were seeded with known cell concentrations of the three pathogenic species, and the limits of detection of the M-PCR tested. The M-PCR was specific, with limits of detection of 10(2)-10(3) cells of the respective species per gram of faeces. CONCLUSIONS: The M-PCR is a rapid, sensitive and specific test for detecting three important enteric bacterial pathogens of pigs. SIGNIFICANCE AND IMPACT OF THE STUDY: The availability of a new diagnostic M-PCR will allow rapid detection and control of three key porcine enteric pathogens.

The distribution of bmpB, a gene encoding a 29.7 kDa lipoprotein with homology to MetQ, in Brachyspira hyodysenteriae and related species.

The distribution of the bmpB gene encoding BmpB, a 29.7 kDa outer membrane lipoprotein of the intestinal spirochaete Brachyspira hyodysenteriae, was investigated. Using PCR, the gene was detected in all the 48 strains of B. hyodysenteriae examined and in Brachyspira innocens strain B256T, but not in 11 other strains of B. innocens nor in 42 strains of other Brachyspira spp. The gene was sequenced from B. innocens strain B256T and from 11 strains of B. hyodysenteriae. The B. hyodysenteriae genes shared 97.9-100% nucleotide sequence similarity and had 97.5-99.5% similarity with the gene of B. innocens strain B256T. Southern hybridisation indicated that bmpB was present on a 1.9 kb HindIII fragment of the B. hyodysenteriae genome and on a 3.1 kb fragment of the B. innocens B256T genome. The B. innocens lipoprotein did not react in Western blots with monoclonal antibody BJL/SH1 that reacts with the B. hyodysenteriae lipoprotein. The difference in binding with the monoclonal antibody may reside in the replacement of a serine residue with a tyrosine residue at base position 210 in the lipoprotein from B. innocens B256T. Comparison of the BmpB amino acid sequence with sequences in the SWISS-PROT protein database indicated that it has 33.9-39.9% similarity with the d-methionine binding proteins (MetQ) of a number of pathogenic bacterial species. The bmpB gene was confirmed to be the same as a gene of B. hyodysenteriae that was recently designated "blpA".

A cross-sectional study to investigate the occurrence and distribution of intestinal spirochaetes (Brachyspira spp.) in three flocks of laying hens.

A cross-sectional study was conducted on a commercial egg-producing farm with a history of wet litter. A total of 600 fresh caecal faecal samples were obtained from under cages of laying hens in three sheds each containing flocks of approximately 5400 hens. Samples were cultured for intestinal spirochaetes, and growth on the primary isolation plate was observed under a phase contrast microscope and subjected to PCRs specific for the intestinal spirochaetes Brachyspira intermedia and Brachyspira pilosicoli. Spirochaete isolates obtained in pure culture were assessed for their ability to cause haemolysis on blood agar and to produce indole, and were typed using pulsed field gel electrophoresis (PFGE). A 1250 base pair portion of the 16S rRNA gene of three B. intermedia and five unidentified isolates was sequenced, and the sequences compared with those of other Brachyspira species. Overall, 121 (20.2%) of the faecal samples contained spirochaetes as determined by growth on the plate and microscopy. Using PCR on the primary growth from these positive samples, 43 (7.2% overall) were shown to contain B. intermedia, 8 (1.3%) to contain B. pilosicoli, and 70 (11.7%) were PCR negative. Only 24 isolates of B. intermedia and five isolates of unknown species were obtained in pure culture. Comparative analysis of the 16S rRNA gene sequence identified the non-B. intermedia isolates as belonging to the proposed species "Brachyspira pulli". PFGE analysis of the B. intermedia strains identified them as having four major banding patterns. Individual patterns were found in hens from different flocks, suggesting cross-transmission of strains between flocks. No environmental sources of infection were identified. The youngest flock had a significantly lower level of colonisation with B. intermedia than the flock of intermediate age (P = 0.004), suggesting that following initial infection of individual young hens on this farm there was amplification and transmission of infection amongst members of the flock.

Survival of intestinal spirochaete strains from chickens in the presence of disinfectants and in faeces held at different temperatures.

This study aimed to evaluate the efficacy of some commonly used disinfectants in inactivating the pathogenic avian intestinal spirochaetes Brachyspira intermedia and Brachyspira pilosicoli, and to examine spirochaete survival in chicken caecal faeces held at 4 degrees C, 25 degrees C or 37 degrees C. Six disinfectants were evaluated at their recommended working concentrations: alkaline salts, quaternary ammonium, iodine as an iodophor, chlorine from a chlorine-release agent, glutaraldehyde and hydrogen peroxide. All but alkaline salts inactivated two different concentrations of both spirochaete species in less than 1 min in the presence of organic matter. Both spirochaete species at three different cell concentrations survived in caecal faeces at 37 degrees C for between 2 and 17 h. B. intermedia tended to survive for longer than B. pilosicoli, but the maximum survival time for both species at 4 degrees C was only 72 to 84 h. Hence, avian intestinal spirochaetes are rapidly inactivated by several common disinfectants, and their survival time in chicken caecal faeces is much less than has been reported for porcine intestinal spirochaetes in porcine faeces. It should be relatively easy to break the cycle of infection between batches of laying birds by resting sheds for a few days, and by using disinfectants on any residual faecal matter.

Dietary enzyme and zinc bacitracin reduce colonisation of layer hens by the intestinal spirochaete Brachyspira intermedia.

Brachyspira intermedia strain HB60 was used to experimentally infect 40 individually caged 22-week-old laying hens. Another 10 control birds were sham-inoculated with sterile broth. All chickens received an experimental layer diet based on wheat. The infected birds were randomly divided into four groups of 10, with the diet for each group containing either 50 ppm zinc bacitracin (ZnB), 100 ppm ZnB, 256 ppm of dietary enzyme (Avizyme), 1302), or no additive. Birds were kept for 6 weeks after infection, and faecal excretion of B. intermedia, faecal water content, egg numbers, egg weights and body weights were recorded weekly. Control birds remained uninfected throughout the experiment. B. intermedia was isolated significantly less frequently from the groups of experimentally infected birds receiving ZnB at 50 ppm or enzyme than from those receiving 100 ppm ZnB or no treatment. Infected birds had a transient increase in faecal water content in the week following challenge, but no other significant production differences were detected amongst the five groups of birds in subsequent weeks. It was not established how the ZnB at 50 ppm and the dietary enzyme reduced the ability of the spirochaete to colonise, but it may have been by bringing about changes in the intestinal microflora and/or the intestinal microenvironment.